1. Field of the Invention
The present invention relates to a fusion protein of PRRS subunit vaccine inducing PRRSV neutralization titers in pigs.
2. Description of Related Art
Porcine reproductive and respiratory syndrome is a porcine infectious disease that primarily strikes respiratory tracts of pigs at various ages and results in sows having reproductive dysfunction. PRRSV is a tough and resistant virus not prone to evoke antibodies having neutralization titers in the infectious pig. Besides, PRRSV is a RNA virus and reproduces easily on the basis of a simplified genetic system, so the probability of genetic mutations is very high. Furthermore, infections and pathogenesis pathway of PRRS virus can be sorted into two stages: (A) infections of epithelium tissues of upper and lower reproductive tracts; and (B) infections of monocytes and macrophages in tissues surrounding reproductive tracts. Thus, the host must have body fluid immunity as well as mucosal immunity with neutralization titers and also a cell-mediated immune response to facilitate removal of the infected viruses and strengthen the host protection mechanisms. However, it is not very easy for PRRSV infectious pig to have neutralization titers in natural conditions, hence typical antibodies basically have little effect on PRRSV, and they even induce mutations in viruses. Furthermore, in antibody-dependent enhancement of phagocytosis, the antibodies could only cause more severe PRRSV infections.
Taiwan Patent No.1-2289933 (also as U.S. patent publication no. 2004/02147617) discloses a target-cell-specific fusion protein, which utilizes a moiety and a functional domain of Pseudomonas aeruginosa exotoxin to fuse a PRRSV ORF7 nuclear protein fragment, with a KDEL signal peptide added on the carboxyl terminus. The fusion protein can be mass-produced in E. coli. When immunized the fusion protein to pigs, it is possible to decrease or to eliminate viremia after being PRRSV-challenged in the immunized pigs. The full text of the patent is incorporated herein.
In heterodimerization between ORF5 and ORF6 of PRRSV, epitope Cys-34 of ORF5 and epitope Cys-8 of ORF6 play a critical role in viral infection and the envelope assembly thereof. (Snijder Eric J., Jessica C. et al., Journal of Virology, January 2003, Vol. 77, No. 1:97-104). Besides, the consensus sequence of PRRSV ORF5 (YKNTHLDLIYNA) is an epitope between amino acid 38th and amino acid 44th, which is located at N-terminal extracellular domain of PRRSV ORF5 and had been identified as a neutralization epitope (Ostrowski M., J. A. Galeota, et al., Journal of Virology, May 2002, Vol. 76, No. 9:4241-4250).
Prior arts disclose constructing whole PRRSV ORF5 or ORF6 antigens between PE and KDEL. After immunization of these fusion proteins, the pigs suffered from severe inflammation in their lungs post being PRRSV-challenged, indicating that PRRSV ORF5 or ORF6 have an antigen-specific allergy effect. Manifestly, it is difficult to use them as PRRS vaccine antigens. Thus, to develop a vaccine and effectively protect pigs from PRRS infections, there are a lot of difficulties that have to be overcome. It should need to be designed such as a lower immunotoxicity and having a high neutralization titer.